摘要
LrrG和表面免疫原性蛋白(Sip)是无乳链球菌(Streptococcus agalactiae)的2种表面蛋白,具有良好的免疫原性。为获得罗非鱼无乳链球菌表面蛋白LrrG和Sip蛋白的融合蛋白,该试验采用基因拼接技术中的双酶切法分2步逐个将Sip和LrrG基因插入pColdⅡ载体中,构建原核表达载体pColdⅡ-LrrG-Sip。将成功构建的融合基因原核表达载体转化感受态细胞BL21(DE3),进行诱导表达条件的优化。结果显示,15℃、IPTG 0.5 mmol·L-1诱导9 h,目的蛋白呈可溶状态的表达量最高。Western Blot检测结果显示LrrG-Sip融合蛋白大小与预测一致(162kDa),说明成功构建了融合基因,为罗非鱼源无乳链球菌亚单位疫苗的研制奠定了基础。
LrrG （leucine-rich repeat protein from GBS） and Sip （surface immunogenic protein）, which are two kinds of surface anti- gen proteins from Streptococcus agalactiae in tilapia, have good immunogenicity. To obtain the LrrG-Sip fusion protein via coalescing surface antigen protein LrrG and Sip of S. agalctiae in tilapia, we cloned Sip and LrrG genes into vector pCold 11 one by one using double enzyme method of gene splicing technology, and constructed a prokaryotic expression vector pCold II -LrrG-Sip. The recombinant plasmid was transformed into E. coli BL21 （DE3） , and the result indicated that 9 h, 15 ℃ , 0. 5 mmol·L^-1 IPTG were the optimum inducing conditions under which fusion protein was most soluble and abundant. Western blotting test showed that the LrrG-Sip fusion protein was about 160 kDa, consistent with the prediction （162 kDa） , which suggested the prokaryotic expression vector pCold II -LrrG-Sip was constructed successfully and laid the foundation for developing subunit vaccines for S. agalctiae in tilapia.
出处
《南方水产科学》
CAS
CSCD
北大核心
2014年第5期17-23,共7页
South China Fisheries Science
基金
现代农业产业技术体系建设专项资金(CARS-49)
广州市科技计划项目(2013J4100078
201300000064)
